The main objective of this research is to investigate the biochemical properties and hormonal regulatory mechanisms of cyclic nucleotide phosphodiesterases. The direction of these studies is to provide an integrated, systematic, and detailed analysis of the function of cyclic nucleotide phosphodiesterases in the processes of growth, differentiation, and hormone action. Linear sucrose gradients, ion-exchange chromatography, isoelectric focusing, and gel filtration are the primary fractional techniques to be used for enzyme separation, identification, and comparison. Cultures of baby hamster kidney fibroblasts (BHK 21 c/13) and purified human peripheral blood lymphocytes will be used to test the hypothesis that activation of specific forms of phosphodiesterase activity involves the mobilization from membranes and/or synthesis of factors as a consequence of interactions of hormones and their cellular receptors. Analytical and preparative techniques will be used to establish the existence of factors produced by hormonal stimulation. Protein synthetic dependent and independent mechanisms of activation of high affinity cyclic nucleotide phosphodiesterases by serum, insulin, methylxanthines, and plant lectins such as concanavalin A and phythohemagglutinin will be studied in these cells. Knowledge gained from these investigations will be applied toward studies of hormone modulation of cyclic nucleotide phosphodiesterase activity in adipose and hepatic tissues. Other potential phosphodiesterase control mechanisms such as the redistribution or translocation of enzymes will be studied during modulation of the growth cycle of cultured 3T3 fibroblasts. In addition, we propose to purify dog kidney cyclic nucleotide phosphodiesterase to homogeneity on a preparative scale and initiate biochemical and immunological characterization of these enzyme forms.